No abstract

Human amnion/chorion tissue derived from the placenta is rich in cytokines and growth factors known to promote wound healing; however, preservation of the biological activities of therapeutic allografts during processing remains a challenge. In this study, PURION® (MiMedx, Marietta, GA) processed dehydrated human amnion/chorion tissue allografts (dHACM, EpiFix®, MiMedx) were evaluated for the presence of growth factors, interleukins (ILs) and tissue inhibitors of metalloproteinases (TIMPs). Enzyme-linked immunosorbent assays (ELISA) were performed on samples of dHACM and showed quantifiable levels of the following growth factors: platelet-derived growth factor-AA (PDGF-AA), PDGF-BB, transforming growth factor α (TGFα), TGFβ1, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), placental growth factor (PLGF) and granulocyte colony-stimulating factor (GCSF). The ELISA assays also confirmed the presence of IL-4, 6, 8 and 10, and TIMP 1, 2 and 4. Moreover, the relative elution of growth factors into saline from the allograft ranged from 4% to 62%, indicating that there are bound and unbound fractions of these compounds within the allograft. dHACM retained biological activities that cause human dermal fibroblast proliferation and migration of human mesenchymal stem cells (MSCs) in vitro. An in vivo mouse model showed that dHACM when tested in a skin flap model caused mesenchymal progenitor cell recruitment to the site of implantation. The results from both the in vitro and in vivo experiments clearly established that dHACM contains one or more soluble factors capable of stimulating MSC migration and recruitment. In summary, PURION® processed dHACM retains its biological activities related to wound healing, including the potential to positively affect four distinct and pivotal physiological processes intimately involved in wound healing: cell proliferation, inflammation, metalloproteinase activity and recruitment of progenitor cells. This suggests a paracrine mechanism of action for dHACM when used for wound healing applications.

The objectives of this study were to determine the viscoelastic shear properties of articular cartilage and to investigate the effects of the alteration of proteoglycan structure on these shear properties. Glycosidase treatments (chondroitinase ABC and Streptomyces hyaluronidase) were used to alter the proteoglycan structure and content of the tissue. The dynamic viscoelastic shear properties of control and treated tissues were measured and statistically compared. Specifically, cylindrical bovine cartilage specimens were subjected to oscillatory shear deformation of small amplitude (gamma degrees = 0.001 radian) over a physiological range of frequencies (0.01-20 Hz) and at various compressive strains (5, 9, 12, and 16%). The dynamic complex shear modulus was calculated from the measurements. The experimental results show that the solid matrix of normal articular cartilage exhibits intrinsic viscoelastic properties in shear over the range of frequencies tested. These viscoelastic shear properties were found to be dependent on compressive strains. Our data also provide significant insights into the structure-function relationships for articular cartilage. Significant correlations were found between the material properties (the magnitude of dynamic shear modulus, the phase shift angle, and the equilibrium compressive modulus), and the biochemical compositions of the cartilage (collagen, proteoglycan, and water contents). The shear modulus was greatly reduced when the proteoglycans were degraded by either chondroitinase ABC or Streptomyces hyaluronidase. The results suggest that the ability of collagen to resist tension elastically provides the stiffness of the cartilage matrix in shear and its elastic energy storage capability. Proteoglycans enmeshed in the collagen matrix inflate the collagen network and induce a tensile prestress in the collagen fibrils. This interaction of the collagen and proteoglycan within the cartilage matrix provides the complex mechanism that allows the tissue to resist shear deformation.

PURIONV R processed dehydrated human amnion/chorion membrane (dHACM; MiMedx Group, Marietta, GA) tissue products were analyzed for the effectiveness of the PURION V R process in retaining the native composition of the amniotic membrane and preserving bioactivity in the resulting products. dHACM was analyzed for extracellular matrix (ECM) composition through histological staining and for growth factor content via multiplex ELISA arrays. Bioactivity was assessed by evaluating endogenous growth factor production by human dermal fibroblasts in response to dHACM and for thermal stability by mechanical tests and in vitro cell proliferation assays. Histology of dHACM demonstrated preservation of the native amnion and chorion layers with intact, nonviable cells, collagen, proteoglycan, and elastic fibers distributed in the individual layers. An array of 36 cytokines known to regulate processes involved in inflammation and wound healing were identified in dHACM. When treated with dHACM extracts, bioactivity was demonstrated through an upregulation of basic fibroblast growth factor, granulocyte colony-stimulating factor, and placental growth factor biosynthesis, three growth factors involved in wound healing, by dermal fibroblasts in vitro. After conditioning at temperatures ranging from 278.7 to 173.5 C, dHACM retained its tensile strength and ability to promote proliferation of dermal fibroblasts in vitro. Elution experiments demonstrated a soluble fraction of growth factors that eluted from the tissue and another fraction sequestered within the matrix. The PURION V R process retains the native composition of ECM and signaling molecules and preserves bioactivity. The array of cytokines preserved in dHACM are in part responsible for its therapeutic efficacy in treating chronic wounds by orchestrating a "symphony of signals" to promote healing.